New polymers and applications

ABSTRACT

The present invention provides a biodegradable, biocompatible polymer that is capable of forming particles (micelles), vesicles, surfaces and membranes, and other structures in which a biologically active agent, e.g. a drug, can be incorporated in such a way that its release to the host can be controlled to a high degree of accuracy, or in where surfaces of the formed polymers can be used to increase the hemeo compatibility of biomaterials. The present invention provides polymer compounds comprising at least one biodegradable polyester having a terminal functional group based on hydrophilic moieties from a phospholipid.

The present invention relates to a novel class of polymers, macroaggregates formed by said polymers, and various uses of said polymersand aggregates for controlled release of substances or to providetemporary coatings to enhance the blood compatibility of biomaterials.

BACKGROUND OF THE INVENTION

Polymers are a versatile class of materials that offer numerous benefitscompared to other material groups. Polymer structures have also beenused to facilitate solutions to a variety of biomedical problems.Remarkable properties such as biocompatibility and/or biodegradability,have been the reasons why they have been used in, for instance, suturesand bioactive membranes. Promising future applications involves areassuch as platforms for tissue regeneration, stent coatings, replacementmaterials for eye lenses and various cosmetic solutions.

Another application is the growing need for new sophisticated and“smart” materials for active drug delivery. Controlled drug deliverytechnology represents one of the most challenging areas of polymerresearch, and the need for new release systems is high. Such deliverysystems offer numerous advantages compared to conventional dosage forms,including improved efficacy, reduced toxicity and improved patientcompliance and convenience. Such systems often use synthetic polymers ascarriers for the drugs. Although the introduction of the first clinicalcontrolled release systems occurred less then 25 years ago, 1997 salesof advanced drug delivery systems in the United States alone wereapproximately $14 billion dollars.

The methods of controlled release are generally divided into twoclasses; temporal control and distribution control. In temporal control,drug delivery systems aim to deliver the drug over an extended durationor at a specific time during treatment. In distribution control, drugdelivery systems aim to target the release of the drug to the precisesite of activity within the body. The two methods have distinctdifferences, and in every situation there is a certain need that can befulfilled depending on the choice of release system.

In order to establish working platforms suitable for either temporal ordistribution control release systems, the use of polymers have beenwidely used. Many polymer classes have been used, including polyesters,polyorthoesters, polyanhydrides, phosphorous containing polymers, andpolyamides. Moreover, numerous examples of hydrophobic/hydrophilic blockcopolymers with surfactant properties have also been made. One exampleis the polylactic acid (PLA) polyethylene glycol (PEG) copolymer systemin which the PEG chain adds hydrophilic properties whereas the PLA chainis hydrophobic, the whole structure being biodegradable. Attempts totarget the degradation of the polymer have been made using combinationsof phosphoesters and aliphatic polyesters, e.g. U.S. Pat. No. 6,166,173.Furthermore, the use of vesicles has provided an alternative releasemethod and the stability of such systems has been increased, e.g. WO99/65 466.

A phenomenon often observed with controlled release formulations ofmedicinal products is that of the “burst effect”, that is, a very largeinitial release of the active substance. In certain cases, this effectmay be desirable. On the other hand, there are cases where it may proveto be dangerous. This is the case, which is particularly detrimental tohormone therapies, which use active principles having very troublesomeor even toxic side effects in high concentrations. In such cases, it isimperative to be able to ensure slow and uniform release in smallquantities of the active principle.

Attempts to overcome such effects have been made, see e.g. U.S. Pat. No.6,319,512. The invention claimed in this patent provides an implant forthe controlled release of at least one pharmaceutically active agent,said implant comprising a core which contains at least one active agentand a sheath which surrounds said core, and is wherein said sheath iscomposed of at least one polymeric film applied around said core.According to a preferred embodiment of that invention, the sheath iscomposed of at least two polymeric films, one surrounding part of thecore and the other surrounding the remaining part. This is however, afairly complex structure, requiring fairly complex manufacturing, andthus has disadvantages in terms of manufacture cost.

Thus, over the last two to three decades there has been a development ofcontrolled release systems for medical use. Numerous patents have beenfiled and granted on such systems. These systems have been based onvarious kinds of structures, such as micelles, vesicles, surface boundagents, etc.

Parallel to this development, the need for materials with new biomedicalfunctions has increased, and new areas in which the use of polymers hasbeen introduced are, for instance, bone implant replacements, stenttechnology, and scaffolds for tissue engineering.

The research groups of Nakabayashi and Ishikara have developed a newtype of copolymer in which a hydrophobic polymer has been used incombination with hydrophilic phosphatidyl choline units. By doing so, anew biocompatible amphiphilic structure was created. To mention a fewcopolymer systems developed by Nakabayashi et al are variouspolymethacrylates, polysulfones, polyethylenes and polystyrenes, whichhave been used in combination with phosphatidyl choline units. Some ofthe most significant improvements compared to the homopolymer have beenincreased blood compatibility and reduced plasma protein adsorption.These effects have been studied in membranes and surfaces as well as inparticles (micelles). It has been shown that the phosphatidyl cholineunit may interact with phospholipids to create stable biomembranes aswell as an ability of the zwitterionic head group to strongly bind waterthereby minimizing the polymer protein interaction leading to increasedhemeo compatibility. Since the first published data was released in theearly 90's, many other research groups have contributed to furtherresearch in the area.

As already indicated, during the last decade polymer research has beendriven towards the design of materials with multiple properties. Thisincludes both new polymerization techniques as well as the use ofpolymers in combination with other highly ordered and controlledstructures. The development of dendritic, hyper branched and star-likestructures parallel to advances in ring-opening metathesis (ROMP), atomtransfer radical (ATRP) and ring-opening (ROP) polymerization techniqueshave enabled the preparation of well-defined functional polymericmaterials with predictable molecular weights and narrowpolydispersities. This development has enabled the synthesis of avariety of new architectures developed from a number of differentbuilding blocks.

Many of these have been proven successful; however, a biodegradablesystem with bio-mimicking and non-thrombogenic properties suitable fordrug release or enhanced blood compatibility has not yet been developed.This would allow materials to be developed with self-regenerating,anti-fouling surfaces with drug releasing capabilities. Furthermore,introducing “phospholipid-like” analogues with different charged ionicgroups would facilitate the combination of specific interactionsprovided by the charged “phospholipid-like” polymer with the biomimeticpolymer that prevent non-specific interaction. Combinations thereofshould promote the binding of charged hydrophilic compounds in additionto the incorporation of hydrophobic water-insoluble ones. In addition,this is in similarity to biological environments where for instance thecell membrane-bound phosphatidyl serine has a negative charge. Moreover,the possibility of drugs delivered from loaded particles of suchmolecules reaching the target cell should increase, thereby facilitatingmore precise and controlled transport and drug release. This shouldreduce unwanted side- and release effects. In addition, the degradationleads to products readily metabolized by the human body. Suchphospholipid analogues could also enhance the stability of liposomesused in drug delivery or be used in combination with naturally occurringphospholipids to create biomembranes. Applications also include cosmeticformulations.

SUMMARY OF THE INVENTION

Thus, in view of the need for new materials suitable for controlledrelease of e.g. medicaments having biodegradable and biocompatibleproperties, the object of the invention is to make available abiodegradable, biocompatible polymer, with increasedblood-compatibility, which is capable of forming particles (micelles),vesicles, surfaces and membranes, and other structures in which abiologically active agent, e.g. a drug, can be incorporated in such away that its release to the host can be controlled to a high degree ofaccuracy.

This object is achieved in a first aspect of the invention by a novelpolymer compound as defined in claim 1.

In a further aspect there is provided a macromolecule in the form of aself-assembled micelle, dendrimer or membrane structure-based on thepolymer defined in claim 1. This macromolecule is defined in claim 2 andclaim 3.

There is also provided, in a third aspect of the invention, a vehiclefor the controlled release of biologically active agents, e.g. drugs,said vehicle being defined in claim 13. Preferred forms of said vehiclein the form of micelles, vesicles, membranes, and surfaces are definedin the claims depending from claim 13.

Finally, there is also provided a method of making the polymer andpolymer aggregates, the method being defined in claims 18-28. The methodmakes it possible to produce polymers being either anionic, cationic orzwitterionic or neutral, or any combination thereof.

The present invention polymers have several advantages for use insystems for controlled drug release or to provide surfaces with enhancedblood compatibility. One advantage is that, in the present invention thepolymers are compatible with blood, a property imparted by thebiomimetic phosphatidyl choline (PC). The polymer is also biodegradable.Moreover, the combination of hydrophilic and hydrophobic segments of thematerial gives the present invention polymers the appropriate physicalproperties needed to form particles or membranes. In addition, the highlevel of synthetic control also leads to control of functionality,thereby increasing the flexibility of this new polymer material, i.e.this material makes it possible to incorporate various types of drugs.With the present invention and the accompanying technique forring-opening polymerization of cyclic esters, it is possible to tailorthe length of the polymers or in a later step the size of particles,depending on the application. Particle or membrane formation can beachieved either by self-assembly of linear polymers, or alternatively,by a dendritic approach in order to form a “one molecule-one particle”type of system.

Further scope of applicability of the present invention will becomeapparent from the detailed description given hereinafter. However, itshould be understood that the detailed description and specificexamples, while indicating preferred embodiments of the invention, aregiven by way of illustration only, since various changes andmodifications within the spirit and scope of the invention will becomeapparent to those skilled in the art from this detailed description.

BRIEF DESCRIPTION OF THE DRAWINGS

The present invention will become more fully understood from thedetailed description given herein below and the accompanying drawingswhich are given by way of illustration only, and thus not limitative ofthe present invention, and wherein

FIG. 1 illustrates the synthetic route for terminated polyε-caprolactone-phosphatidyl choline (PCL-PC) according to the presentinvention.

FIG. 2 illustrates micelle formation of an amphiphilic moleculeaccording to the present invention.

FIG. 3 shows an example of a dendrimer structure, a branchedpolyfunctional one particle molecule, according to the present invention

FIG. 4 exemplifies other cyclic esters that, in addition toε-caprolactone, that could be used to synthesise the polymer accordingto the present invention.

FIG. 5 shows a schematic model of possible molecular arrangement in aPCL-PC blend in the form of cast films (left) and after heat treatmentin water (right).

FIG. 6 shows a diagram depicting formation of TAT-complex when using aPCL-PC-material in contact with whole blood.

DETAILED DESCRIPTION OF PREFERRED EMBODIMENTS

The approach has been to combine the use of phosholipid moieties incombination with biodegradable polyesters in order to prepare a fullybiocompatible and biodegradable polymer system. One major goal has beento design macromolecules so they form a certain type of structuredepending on the application. Two examples are membranes and micelles.

The present invention provides polymer compounds comprising at least onebiodegradable polyester having a terminal functional group based on thehydrophilic moiety in phospholipid.

The polymer compounds according to the present invention can beaggregated and have the shape of micelles, vesicles and membranes. Thepolymer compounds can also be designed such that they emanate from acentral core so as to form a dehdrimer. The dendrimer-type of polymercompound forms an essentially spherical particle with said functionalgroups forming the surface layer of said spherical particle or isconcentrated at the surface, thus mimicking the surface of vesicles.

A solution of the micelles or spherical particles formed by the polymercompound according to the present invention can be used as a drugformulation, where the micelles or particles enclose a medicament.

The polymer compound according to the present invention can further beused for coating an object, e.g. a vehicle, and the thus formed coatingmay be loaded with an (biologically) active agent, e.g. a drug. Thecoating constitutes a layer having a thickness of 0.1-100 μm, saidfunctional groups forming an outer layer of said coating.

The coated object may be used in biological or medical applications,such as a medical device, medical device for implantation, stent,artificial orthopedic device, spinal implant, joint implant, attachmentelement, bone nail, bone screw, or a bone reinforcement plate.

The biodegradeable polyester used in the polymer compound according tothe present invention is polymerized from a cyclic monomer selected fromthe group of cyclic esters and carbonates including ε-caprolactone,lactide, glycolide, β-butyrolactone, propiolactone,trimethylenecarbonate and combinations thereof.

The terminal functional group of the polymer compound according to thepresent invention is positively or negatively charged, or iszwitterionic or electrically neutral. The terminal functional group isselected from but not restricted to phosphatidyl choline, phosphatidylethanolamine, phosphatidyl serine, ammonium salt, carboxylic acid orcarboxylate, phosphoric acid, phosphate, phosphonate, sulphonate,sulphonic acid, peptide, nucleotide, carbohydrate.

The molecular weight of the polymer compound according to the presentinvention can be in the range 1000-200 000 g/mol, preferably 20 000g/mol. The present invention also provides a method of preparing abiodegradable and biocompatible polyester having a terminal functionalgroup based on a phospholipid, which is comprised by the followingsteps: reacting a cyclic ester monomer and an alcohol in the presence ofa catalyst/an initiator to provide a ring opened polymer having an —OHterminal end; reacting the —OH terminal end of the obtained polymer witha phosphorous containing compound to provide a polymer having aphosphate terminated polymer; and reacting said phosphate terminated endof said polymer to obtain a polymer having functionalized end.

The phosphorous containing compound in said method is preferablyselected from the group consisting of ethylene chloro phosphates. Insaid method, the step of providing a functionalized polymer alsocomprises reacting the terminal end with trimethylamine. The resultingpolyester is preferably poly ε-caprolactone-phosphatidyl choline.

The present invention further provides a method of preparingbiodegradable and biocompatible polyester amphiphiles having a chargedterminal functional group in combination with phosphatidyl choline, themethod comprising the following steps: reacting a cyclic ester monomerand an alcohol in the presence of a catalyst/initiator to provide aring-opened polymer having an —OH terminal end; and reacting said —OHterminal end of the obtained polymer with a co-halo acid halide toobtain an alkyl halide; and reacting said polymer/polymers to obtain apolymer having a functionalized end.

In said method, the step of providing a functionalized polymer comprisesreacting the terminal end with trimethylamine. The resulting polyesteris preferably poly ε-caprolactone-ammonium salt.

The present invention further provides a method of preparing abiodegradable and biocompatible polyester amphiphiles having a chargedterminal functional end in combination with phosphatidyl choline,comprising the steps of reacting a cyclic ester monomer and an alcoholin the presence of a catalyst/an initiator to provide a polymer havingan —OH terminal end; reacting the —OH terminal end of the obtainedring-opened polymer with a succinic anhydride to produce afunctionalized (carboxylic acid)- or carboxylate-terminated polymer.

In said method, the step of providing a functionalized polymer comprisesreacting the terminal end with derivatives of carboxylic acid or itsanhydrides. The resulting polyester is preferably polyε-caprolactone-carboxylic acid or poly ε-caprolactone-carboxylate.

Now the general experimental methods will be described.

Tin(II)triftuoromethane sulfonate (Sn(OTf)₂) was purchased from Aldrichand was azeotropically distilled with toluene prior to use.ε-caprolactone (ε-CL) and triethylamine were purchased from Aldrich andwere distilled over calcium hydride prior to use. Chloroform anddichloromethane (VWR) were washed over a basic aluminum oxide (Al₂O₃)column and distilled over CaH₂ prior to use. Succinic anhydride(Aldrich) was recrystallized from dry chloroform and stored in a glovebox prior to use. 4-chlorobutyrylchloride (Aldrich) was used asreceived. Acetonitrile was purchased from Lancaster and was distilledfrom magnesium sulfate prior to use. Ethylene chloro phosphate waspurchased from Lancaster and was distilled and stored in a freezer priorto use. Benzyl alcohol was purchased from Aldrich and was distilled overcalciumhydride prior to use. ¹H-NMR and ³¹P-NMR were performed on a JEOL400 MHz. SEC was performed on a Waters instrument.

The following section will be based on FIG. 1, which illustrates thesynthetic route for terminated poly ε-caprolactone-phosphatidyl choline.

Synthesis of Poly ε-Caprolactone. PCL (Step I in FIG. 1.)

A 50 ml two necked Schlenk flask was added a stir bar and the flask wassealed with a septum. The equipped flask was carefully flame-dried undervacuum and purged with nitrogen. For the polymerization theε-caprolactone monomer (10.0 g, 87.6 mmol) and the Sn(OTf)₂ catalyst(0.063 g, 0.11 mmol), using 5 mol % to initiator, were added in a glovebox. Following removal of the flask the initiator benzyl alcohol (0.23g, 2.2 mmol for a degree of polymerisation of 40) was syringed into theflask under protecting gas. The mixture was stirred vigorously andrapidly heated to 110° C. Following completion of the reaction (t=60minutes), the polycaprolactone (PCL) mixture was dissolved in THF andprecipitated in 500 ml of cold methanol. The precipitate was filteredand washed repeatedly with methanol and then dried in vacuum at 40° C.until a constant weight was reached.

Synthesis of PCL Coupled to Ethylene Chloro Phosphate (Step II in FIG.1)

For the phosphorylation 4.0 g (0.86 mmol) of a PCL with a degree ofpolymerisation (DP) of 40 was weighed in a pre-dried nitrogen flask anddissolved in 20 ml dry dichloromethane (CH₂Cl₂). 1.5 equivalents of drypyridine (0.11 ml, 1.29 mmol) were thereafter added under nitrogen. Theflask was attached to a pre dried dropping funnel and attached to anitrogen inlet and thereafter cooled to −5° C. 5 mL of dry CH₂Cl₂ and 2equivalents of ethylene chloro phosphate (0.14 g, 1.028 mmol) was addedthe dropping funnel. The solution was slowly added drop wise and thesolution was stirred for approximately 2 hours and then slowly allowedto reach ambient temperature and further stirring for 4 hours. After thereaction was completed the solution was charged with an additional 50 mlof CH₂Cl₂ and extracted twice with distilled water (50 ml) and twicewith a 1 M NaHCO₃ (50 ml) solution to remove the, from the reactionformed, pyridinium salt and excess ethylene chloro phosphate reagent.Thereafter the organic phase was separated and dried from water usingsodium sulphate, stirring for 30 minutes. 50 ml of toluene was added andthe organic phases as well as trace amounts of pyridine was removed byrotational evaporation at ambient temperature.

Synthesis for the Ring-Opening of Ethylene Phosphate to YieldPhosphatidyl Choline Terminated PCL (Step III in FIG. 1)

For the formation of PC terminated PCL 1.0 g (0.21 mmol) of 2 wasweighed in a 50 mL pre-dried round-bottom flask and thereafter dissolvedin 10 ml of dry acetonitrile. The solution was transferred to a pressuretube with two stopcocks, purged with nitrogen and sealed, thereaftercooled to −10° C. Two equivalents (0.42 mmol, 39 μl) of trimethylaminewto the PCL polymer was carefully condensed into the pressure tube andthereafter slowly heated to 60° C. The pressure tube was left understirring for 45 hours and then left to cool to ambient temperature andthe reaction product was precipitated in cold methanol. The precipitatewas collected and dried until constant weight.

Synthesis of PCL Coupled to Succinic Anhydride

For the synthesis 2.0 g (0.44 mmol) of 1 and 88 mg (0.88 mmol) ofsuccinic anhydride was added a 50 ml pre dried two-necked round bottomflask equipped with a stir bar and purged with nitrogen. The compoundswere dissolved in 15 ml of dry chloroform and a dropping funnel wasattached and the solution cooled to 0° C. 86 mg (0.88 mmol) of triethylamine was added 5 ml of dry chloroform and charged in the funneland slowly added drop wise to the cooled solution under a 30-minuteperiod. The solution was slowly allowed to reach ambient temperature andleft under stirring for an additional 3 hours. Following completeconversion the polymer was precipitated in cold methanol filtrated anddried until constant weight.

Synthesis of PCL with a Terminal Quaternary Ammonium

For the synthesis 2.0 g (0.44 mmol) of 1 and 87 mg (1.10 mmol) ofpyridine was added a 50 ml pre dried two-necked round bottom flaskequipped with a stir bar and purged with nitrogen. The compounds weredissolved in 15 ml of dry chloroform and a dropping funnel was attachedand the solution cooled to −10° C. 116 mg (1.10 mmol) of 4-chlorobutyrylchloride was added 5 ml of dry chloroform and charged in the funnel andslowly added drop wise to the cooled solution under a 30-minute period.The solution was slowly allowed to reach ambient temperature and leftunder stirring for an additional 3 hours. Following complete conversionthe polymer was precipitated in cold methanol filtrated and dried untilconstant weight. The precipitate was thereafter dissolved in 10 ml ofdry acetonitrile. The solution was transferred to a pressure tube withtwo stopcocks, purged with nitrogen and sealed, thereafter cooled to−10° C. Two equivalents (0.42 mmol, 39 μl) of trimethylamine_((g)) tothe PCL polymer was carefully condensed into the pressure tube andthereafter slowly heated to 60° C. The pressure tube was left understirring for 45 hours and then left to cool to ambient temperature. Theformed compound was precipitated in cold methanol and the precipitatewas collected and dried until constant weight.

Results

The emphasis was to synthesise a fully biodegradable polymer incombination with phosphatidyl choline as a possible future carrier forcontrolled drug release or a temporarily coating for enhanced bloodcompatability or other biomedical applications. The ambition has been tointroduce the use of phospholipid analogues into new areas of polymerresearch, keeping in mind what already has been done in this area. Withthe recent development of polymerisation techniques for the synthesis ofbiodegradable polyesters it is not until now that this has beenpossible. Controlled ring-opening polymerisation of for instancelactides and ε-CL now makes it possible to design polyesters withcontrolled molecular weight and narrow polydispersities. It should alsobe pointed out that by the approval from the food and drugadministration (FDA), both PCL and PLA are classified as biocompatiblepolymers, which degrades into molecules acceptable in the humanmetabolism.

Synthesis:

In our initial results a series of various linear PCL with variousmolecular weights was made, mainly to demonstrate the high level ofcontrol in this synthetic route, but also to create the firstamphiphiles with particle or membrane forming properties. The followingtable recall some initial polymerisation data. TABLE 1 Ring-openingpolymerisation data of PCL. Temp. Time Yield I/M Sample CatalystInitiator [° C.] [hrs] [%] Ratio DP PDI 1 Sn(OTf)₂ EtOH 35 4 70 5 5 1.142 Sn(OTf)₂ EtOH 35 15 90 15 17 1.18 3 Sn(OTf)₂ EtOH 35 20 95 30 27 1.074 Sn(OTf)₂ EtOH 35 39 97 60 66 1.19

From Table 1 it is clear that the molecular weight of PCL can becontrolled by the ratio of initiator to monomer, as previouslyexplained. Using ¹H-NMR analysis the PCL was fully characterized andboth the α- and the ω-end groups were identified. The following chemicalshifts were observed for the PCL molecule initiated from benzyl alcohol:¹H-NMR (CDCl₃) δ=1.35 (m, 2H, —CH₂—, poly), 1.65 (m, 2H, —CH₂—, poly),1.65 (m, 2H, —CH₂—, poly), 2.30 (t, 2H, —CH₂—, poly), 3.63 (q, 2H,—CH₂—, ω-end), 4.04 (t, 2H, —CH₂—, poly), 5.10 (s, 2H, —CH₂—, α-end),7.34 (m, 5H, —ArH, α-end)

¹H-NMR analysis could be used to monitor the transformation of hydroxylto ethylene phosphate as the proton group adjacent the hydroxyl group at3.62 ppm was diminished, while at the same time a build up ofresonance's from the ethylene protons in the phosphate was observed at4.32 ppm. Furthermore, ³¹P-NMR analysis provided a second spectroscopyanalysis to track the formation of the ethylene phosphate terminated PCLas the ³¹P-NMR signal of the starting material was shifted from 23.1 ppmto 18.0 ppm in the case of ethylene phosphate. ¹H-NMR (CDCl₃) δ=1.35 (m,2H, —CH₂—, poly), 1.63 (m, 2H, —CH₂—, poly), 1.63 (m, 2H, —CH₂—, poly),2.30 (t, 2H, —CH₂—, poly), 4.04 (t, 2H, —CH₂—, poly), 4.32-4.48 (m, 4H,—CH₂—CH2—, ω-end), 5.10 (s, 2H, —CH₂—, α-end), 7.34 (m, 5H, —ArH, α-end)³¹P-NMR (CDCl₃) δ=18.3

In the last ring-opening step, the final PCL phospatidyl cholinemolecule was also characterised with ¹H-NMR. A distinct singlet from themethylene signals in the choline unit was observed at 3.42 ppm. Theethylene protons in the phosphatidyl unit are now separated at 3.75 ppmand at 4.20 ppm. ³¹P-NMR analysis revealed the phosphorous signal fromthe PC group at −1.1 ppm compared to the intermediate phosphorous signalat 18.0 ppm. From the ¹H- and ³¹P-NMR results, it is clear that thesynthetic route is functioning. Importantly the synthesis could beperformed with complete conversion between each step and with highyields, typically around 90% of PCL-PC.

Having established a synthetic route for the “phospholipid-like” PCL-PCpolymer the scope of the synthesis was enhanced to also include charged“phospholipid-like” polymers having a net anionic or cationic charge. Toprovide a phospholipid analogue with a net anionic charge a PCL with aterminal hydroxyl group was reacted with succinic an hydride in thepresence of triethyl amine resulting in the wanted terminal succinicacid monoester. ¹H-NMR analysis was used to monitor the conversion and abuild up of the succinate protons, i.e. two triplets, were formed at2.65 ppm whereas the ethylene protons adjacent the hydroxyl was shiftedto 4.12 ppm.

¹H-NMR (CDCl₃) δ=1.35 (m, 2H, —CH₂—, poly), 1.65 (m, 2H, —CH₂—, poly),1.65 (m, 2H, —CH₂—, poly), 2.30 (t, 2H, —CH₂—, poly), 2.62 (t, 2H,—CH₂—, ω-end), 2.62 (t, 2H, —CH₂—, ω-end), 4.04 (t, 2H, —CH₂—, poly),5.10 (s, 2H, —CH₂—, α-end), 7.34 (m, 5H, —ArH, α-end)

For the formation of a cationic phospholipid analogue the synthesis wassomewhat more complex and consists of two separate steps. In the firststep 4-chlorobutyryl chloride was reacted with the terminal hydroxylgroup of the polymer. Following purification the intermediate wasredissolved in acetonitrile and reacted with Me₃N at 60° C. to allowformation of the cationic quaternary ammonium salt with the chloride ionas gegen ion. ¹H-NMR was used to characterize the obtained product andthe methyl resonance of the quaternary ammonium salt was observed at3.43 ppm. Furthermore, the proton group adjacent the quatemary ammoniumwas observed at 3.72 ppm.

¹H-NMR (CDCl₃) δ=1.35 (m, 2H, —CH₂—, poly), 1.65 (m, 2H, —CH₂—, poly),1.65 (m, 2H, —CH₂—, poly), 2.10 (m, 2H, —CH₂—, ω-end), 2.30 (t, 2H,—CH₂—, poly), 2.50 (t, 2H, —CH₂—, ω-end), 3.43 (s, 9H, —CH₃ , ω-end),3.72 (t, 2H, —CH₂—, ω-end), 4.04 (t, 2H, —CH₂—, poly), 5.10 (s, 2H,—CH₂—, α-end), 7.34 (m, 5H, —ArH, α-end)

Particle Formation:

Following the synthesis two particle formation experiments wereperformed, mainly to get an indication on how these structures behaved.Particle formation using two different routes was conducted.

Using the first route, a phosphatidyl choline terminated PCL (DP=16) wasdissolved in chloroform (CHCl₃). Thereafter the dissolved compound wasadded drop wise to water. Well-defined drops were formed (two-phasesystem). Following addition, a stir bar was added and stirring wasapplied for approximately 30 minutes, creating a fine-dispersed particlesolution. In the early stage, after the stirring had been stopped,flocculation was observed. However, after 30 minutes of stirring onlystable particles were obtained. By “stable” it is meant that no visualflocculation occurred, indicating stable particles.Environmental-Scanning Electron Microscopy (E-SEM) analysis indicatedparticles with a diameter of 1-10 μm. Evaporation of the chloroformsolidified the particles.

Using the second route, a solvent combination was chosen that couldallow a single phase of a combination of the solvents but with thePCL-phosphatidyl choline being totally soluble in one. An acetone-watercombination was chosen (5 mL/95 mL) and a small amount (10 mg) of thePCL (DP=16)-phosphatidyl choline. At first the compound was dissolved inacetone, and was thereafter added drop wise into water. After theaddition, the solution was perfectly transparent, indicating particlesizes in the nanometer (nm) range.

These two experiments gave an indication on that particle formation ispossible and that the system is surface-active. The desired effect isseen in FIG. 2, which shows the micelle formation of the amphiphilicmolecule.

In FIG. 2 the rings represent hydrophilic phosphatidyl choline unitswhereas the zigzag lines represent hydrophobic PCL chains. The figureschematically shows the self-assembly of these molecules in an aqueousmedium (please observe that the rings could mean an end group which isnot phosphatidyl choline, i.e. anionic or cationic in combination withPC).

Film Properties:

The mechanism explaining the low protein adsorption and cell attachmentfor non-degradable phosphatidylcholine functional polymers involves bothsurface-enrichment of the phosphatidylcholine unit and the attraction ofphospholipids to this surface to form a biomembrane-like structure. Itis therefore conceivable that the biodegradable amphiphile PCL-PC couldfunction the same way in addition to being biodegradable.

Films of the above PCL-PC oligomers were not stable in water. PCL(M_(w)˜80 000 g/mol) blend with PCL-PC were good film-formers and couldbe cast into homogenous films. The surface composition of as cast filmsof PCL/PCL-PC (DP=45) was quite similar to pure PCL as shown by XPS witha C/O ratio of 73/27 with no signs of phosphor or nitrogen. Contactangle of cast PCL/PC films was 65 degrees, which is only slightly lowerthan the 69 degrees measured on pure PCL. This is not surprising,considering the low content of phosphatidylcholine end groups and thehydrophobic nature of PCL. As the system strives to minimize itsinterfacial energy, the phosphatidylcholine chain ends will be buried inthe bulk exposing pure PCL to the polymer-air interface.

Under water, however, interfacial free energy is minimized when thehydrophilic phosphatidylcholine would be enriched at the polymer-waterinterface. Therefore, the PCL/PCL-PC blend film was quickly immersedinto heated water at 90° C. (which is above the melting temperature forPCL) to provide molecular mobility for migration. The film first becametransparent due to melting of the crystalline PCL. Prior to cooling, thefilm again became opaque due to water uptake by micellar domains ofphosphatidylcholine in the bulk at 90° C. During cooling furtheropaqueness occurred as the polymer recrystallised.

Migration of PCL-PC oligomers to the surface was indeed confirmed bycontact angle measurements. The contact angle (advancing) was decreasedto 40 degrees, which is indicative of enrichment of polar groups at thesurface towards water. ESCA spectrum shown in FIG. 5 reveals theappearance of both nitrogen, N1s 2.4%, and of phosphorous, P2p 1.5%arising from the polar, surface-oriented, phosphatidylcholine. Thetheoretical concentration in pure PCL-PC (DP=45) is only 0.3%.

It is likely that some surface rearrangement still occurs in theamorphous top layer when the samples are dried prior to ESCA and contactangle measurements. Surface dynamics should therefore be investigatedfurther using dynamic contact angle. The overall mechanism is summarizedin FIG. 5 where PCL is oriented towards the air surface with PC formingmicellar domains in the bulk of cast films. Upon heating in water,however, the surface rearranges to drive PC towards the polymer-waterinterface.

Whole Blood Measurements:

Since the fate of biomaterials is strongly dependent on the activationof the blood plasma cascade system, like the coagulation system, theformation of thrombin-anti-thrombin (TAT) complexes were studied in theslide chamber model. The slide chamber methodology facilitates in vitroanalysis of biomaterial surfaces in contact with whole blood. A PCL-PCsystem with a DP of 45 was used as well as two reference surfacesconsisting of PCL and polyvinylchloride (PVC). A diagram depicting TATformation is shown in FIG. 6. This result indicates that the PCL-PCsystem holds non-thrombogenic properties and that the formation of TATis largely reduced compared to both PCL and PVC, a well-knownbiomaterial. This effect can be explained by the enrichment of PC groupson the polar surface, which decreases adherence of proteins. Moreover,the thrombocyte count was larger for whole blood in contact with thePCL-PC surface than the PVC reference

Molecular Variation:

To extend the use of this synthetic route, one can also includenon-linear type of molecules. In a totally branched system, e.g.initiated from a polyol or macro initiator, one could obtain a“one-molecule-one-particle” system, in which the self-assembly from manymolecules has been changed into a “one-molecule-one-particle” formingsystem with a controlled size. Dendritic type of structures could forinstance be synthesized from the coupling of benzylidine protectedbis(hydroxymethyl) propionic acid (bis-MPA) with benzyl protectedbis-MPA followed by selective deprotection to yield a first generationdendrimer. An alternative approach would be the tailoring of a dendriticstructure from benzylidene-protected glycerol and 2-bromopropionic acid.The addition of branching points in combination with ring-openingpolymerization gives an almost unlimited source of architecturalpossibilities, and the common link is that the functionality on thesurface will be much higher compared to linear structures. Thehydrophobic unit would still be biodegradable polyester, and thephopshatidyl choline unit imparting hydrophilic properties. One changeis the architecture of the molecule, i.e. the branching points, whichyield a molecule with a much higher functionality on the surface. Theend-functionality, however, does not always have to be phosphatidylcholine; other functionalities or combinations of functionalities canalso be chosen, to add specific interactions, as well as the addition offor example receptor ligands. With this synthetic approach, structure,size and functionality can be controlled. One visual example of such astructure is a branched polyfunctional one particle molecule, as seen inFIG. 3 (please observe that the rings could mean an end group which isnot phosphatidyl choline).

The monomer used in the previously described synthetic routes has in allcases been ε-CL. Recently, the controlled ring-opening polymerization ofother cyclic esters has been investigated, and it is now possible totailor the molecular weight of other polyesters as well. A summary ofother cyclic esters and carbonates, which either separately or incombination, could be used in the described synthesis, is shown in FIG.4. In all cases the, obtained polyester is biodegradable.

Thus, according to the present invention, a fully biodegradablepolyester-phosphatidyl choline compound was synthesized using highlydeveloped polymerization techniques. This molecule had amphiphilicbehavior due to hydrophobic properties from the PCL chain andhydrophilic properties from the phosphatidyl choline unit. PCL is oneexample of a biodegradable polyester, but according to the presentinvention other monomers, such as lactides, could also be used toproduce similar structures. In the present invention synthetic route,only a linear type of molecules was created, but it is also possible toprovide branched/dendritic type of structures with a much higherfunctionality on the surface. The polymers according to the presentinvention can suitably be used in biological and medical applications,for instance as membranes and as drug delivery vectors.

With the invention being thus described, it will be obvious that thesame may be varied in many ways. Such variations are not to be regardedas a departure from the spirit and scope of the invention, and all suchmodifications as would be obvious to one skilled in the art are intendedto be included within the scope of the following claims.

1. A polymer compound, comprising at least one biodegradable polyesterhaving a terminal functional group based on hydrophilic moieties ofphospholipids.
 2. A polymer compound as claimed in claim 1, comprising aplurality of biodegradable polymers emanating from a central core so asto form a dendrimer.
 3. Aggregate of polymers as claimed in claim 1,having the shape of micelles, vesicles and membranes.
 4. A polymercompound as claimed in claim 1, wherein said polyester is polymerizedfrom a cyclic monomer.
 5. A polymer compound as claimed in claim 4,wherein said cyclic monomer is selected from the group of cyclic estersand carbonates.
 6. A polymer compound as claimed in claim 5, whereinsaid cyclic esters and carbonates are selected from the group consistingof ε-caprolactone, lactide, glycolide, β-butyrolactone, propiolactone,trimethylenecarbonate and combinations thereof.
 7. A polymer compound asclaimed in claim 1, wherein the terminal functional group isphosphatidyl choline, phosphatidyl ethanolamine, phosphatidyl serine,ammonium salt, carboxylic acid or carboxylate, phosphonic acid,phosphate, phosphonate, sulphonate, sulphonic acid, peptide, nucleotide,carbohydrate.
 8. A polymer compound as claimed in claim 1, wherein theterminal functional group is positively charged.
 9. A polymer compoundas claimed in claim 1, wherein the terminal functional group isnegatively charged.
 10. A polymer compound as claimed in claim 1,wherein the terminal functional group is zwitterionic or electricallyneutral.
 11. A polymer compound as claimed in claim 1, the molecularweight of which is in the range of 1000-200 000 g/mol, preferably 20 000g/mol.
 12. A dendrimer type polymer compound as claimed in claim 2,forming an essentially spherical particle with said functional groupsforming the surface layer of said spherical particle.
 13. An objectprovided with a coating made of a polymer compound as claimed in claim1, wherein said polymer compound forms a layer having a thickness of0.1-100 μm, said functional groups forming an outer layer of saidcoating.
 14. The object as claimed in claim 13, wherein said coating isloaded with an (biologically) active agent.
 15. The object as claimed inclaim 13, wherein the object is an object used in biological or medicalapplications.
 16. The object as claimed in claim 15, wherein it is amedical device, medical device for implantation, stent, artificialorthopedic device, spinal implant, joint implant, attachment element,bone nail, bone screw, or a bone reinforcement plate.
 17. A drugformulation, comprising a solution of micelles or spherical particlesformed by a polymer compound as claimed in claim 1, wherein the micellesor particles enclose a medicament.
 18. A method of preparing abiodegradable and biocompatible polyester having a terminal functionalgroup based on a phospholipid, the method comprising the followingsteps: reacting a cyclic ester monomer and an alcohol in the presence ofa catalyst/an initiator to provide a ring opened polymer having an —OHterminal end; reacting the —OH terminal end of the obtained polymer witha phosphorous-containing compound to provide a polymer having aphosphate terminated polymer; and reacting said phosphate terminated endof said polymer to obtain a polymer having functionalized end.
 19. Themethod as claimed in claim 18, wherein said phosphorous containingcompound is selected from the group consisting of ethylene chlorophosphate.
 20. The method as claimed in claim 18, wherein the step ofproviding a functionalized polymer comprises reacting the terminal endwith Me₃N.
 21. The method as claimed in claim 18, wherein the resultingpolyester is poly ε-caprolactone-phosphatidyl choline.
 22. The method asclaimed in claim 21, wherein the resulting yield of the polyE-caprolactone-phosphatidyl choline is at least 90%.
 23. A method ofpreparing biodegradable and biocompatible polyester phospholipid-likeanalogues having a cationic terminal functional group, the methodcomprising the following steps: reacting a cyclic ester monomer and analcohol in the presence of a catalyst/an initiator to provide aring-opened polymer having an —OH terminal end; reacting said —OHterminal end of the obtained polymer with a ω-halo acid halide to obtainan alkyl halide; and reacting said polymer/polymers to obtain a polymerhaving a functionalized end.
 24. The method as claimed in claim 23,wherein the step of providing a functionalized polymer comprisesreacting the terminal end with Me₃N.
 25. The method as claimed in claim23, wherein the resulting polyester is poly ε-caprolactone-ammoniumsalt.
 26. A method of preparing a biodegradable and biocompatiblepolyester phospholipid-like analogues having an anionic terminalfunctional group, the method comprising the following steps: reacting acyclic ester monomer and an alcohol in the presence of a catalyst/aninitiator to provide a polymer having an —OH terminal end; and reactingthe —OH terminal end of the obtained ring-opened polymer with a succinicanhydride to produce a functionalized (carboxylic acid)- orcarboxylate-terminated polymer.
 27. The method as claimed in claim 26,wherein the step of providing a functionalized polymer comprisesreacting the terminal end with derivatives of derivatives of carboxylicacid or its anhydrides.
 28. The method as claimed in claim 26, whereinthe resulting polyester is poly ε-caprolactone-carboxylic acid or polyε-caprolactone-carboxylate.